Analysis of Chromatin Opening in Heterochromatic Non-Small Cell Lung Cancer Tumor-Initiating Cells in Relation to DNA-Damaging Antitumor Treatment

• Purpose: We previously reported that sphere-forming non-small cell lung cancer (NSCLC) tumor-initiating cells (TICs) have an altered activation of DNA damage response-and repair proteins and are refractory to DNA-damaging treatments. We analyzed whether chromatin organization plays a role in the observed refractoriness.Methods and Materials: Bulk cells and TICs from the NSCLC H23 and H1299 cell lines were examined using cell viability, clonogenic survival, Western blot, short interfering RNA analysis, and micronucleus assay.Results: NSCLC TICs displayed elevated heterochromatin markers trimethylated lysine 9 of histone H3 and heterochromatin protein 1 gamma relative to bulk cells and reduced cell viability upon histone deacetylase inhibition (HDACi). Vorinostat and trichostatin A increased the euchromatin markers acetylated lysine 9/14 of histone H3 and lysine 8 of histone H4, and HDACi pretreatment increased the phosphorylation of the DNA damage response proteins ataxia telangiectasia mutated and DNA-dependent protein kinase, catalytic subunit, upon irradiation in TICs. HDACi sensitized TICs to cisplatin and to some extent to ionizing irradiation. The protectiveness of a dense chromatin structure was indicated by an enhanced frequency of micronuclei in TICs following irradiation, after knockdown of heterochromatin protein 1 gamma.Conclusions: Although confirmatory studies in additional NSCLC model systems and with respect to analyses of other DNA damage response proteins are needed, our data point toward a heterochromatic structure of NSCLC TICs, such that HDACi can sensitize TICs to DNA damage.

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MEDICIN OCH HÄLSOVETENSKAP  -- Klinisk medicin -- Cancer och onkologi (hsv//swe)

MEDICAL AND HEALTH SCIENCES  -- Clinical Medicine -- Cancer and Oncology (hsv//eng)

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